Phagocytosis is a phylogenetically ancient response to particulate stimuli adapted by specialized cells of the immune system. Among the best characterized phagocytic receptors are those that recognize the Fc portion of IgG (FCyRs). Phagocytosis via FCyRs requires the recruitment and activation of Syk, a tyrosine kinase expressed predominantly in hematopoietic cells. In this proposal we outline approaches to identify and characterize downstream targets of Syk that initiate or modify phagocytic signaling. Using proteomics to probe the "phosphotyrosome" of phagocytosis, we have identified proteins that either undergo enhanced tyrosine phosphorylation, or associate with phosphotyrosyl-containing proteins, during phagocytosis. Among these proteins are moesin, a member of the ERM family of actin-binding proteins and Abpl, an adaptor for F-actin, and dynamin. We hypothesize that moesin and ezrin, 2 ERM family members, stabilize the association of the actin based cytoskeleton with the plasma membrane in nascent pseudopods, thereby contributing to pseudopod rigidity. As both proteins contain functional immunoreceptor tyrosine-based activation motifs (ITAMs) that potentially bind Syk, they may act as direct transducers of the phagocytic signal. In Specific Aim I, we outline approaches to test this hypothesis. These experiments include use of software we helped design to predict effective siRNA sequences. In Specific Aim II, we will determine whether Abpl and/or its family members, cortactin and HS1, coordinate the delivery of cytoskeletal components with endo-/exocytic machinery (dynamin). As we have previously proposed a "two compartment" model of phagocytosis whereby the programs of cytoskeletal assembly and membrane trafficking both contribute to phagocytosis, Abp1 and its homologs would fulfill the role of molecular coordinator of these events.